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Nov 15, 2006 (Vol. 26, No. 20)

Filtration Technology Steps Up to Next Level

Bioprocessors' Evolving Separation Needs Continue to Fuel Innovative Approaches

  • Process streams look very different from five or ten years ago, states Jean Paul Mangeolle, president of Millipore’s (www.millipore.com) bioprocess division. Rising protein titers have caused many processes to shrink, but more concentrated process fluids strain downstream operations designed for more dilute process streams. “Yesterday’s 20,000-liter tank is now a 5,000-liter tank, and it most likely incorporates some type of disposable technology,” says Mangeolle.

    Not too long ago, sterilization of filters, stainless steel housings, and validating the cleaning process were the rules. Today, disposable filtration, tubing, connectors, and storage vessels have become standard. “We can now perform multiple transfers of protein products between containers without having to use a single reusable filter,” says Mangeolle. Disposable filters have been a boon to monoclonal antibody production, which is filter- and separation-intensive and relies heavily on clarification and viral clearance.

    Interest in vaccines, particularly from emerging vaccine cell culture processes, will stimulate interest in filtration products for cell culture as well as virus harvesting. Vaccine developers are close to commercializing cell-culture influenza vaccines that will reduce reliance on chicken eggs as a source for yearly flu shots.

    Since the disposable revolution began about ten years ago, the filtration business has evolved from its former pure-play product focus to more of a solutions model. Filtration customers expect not just a membrane but a disposable housing; a way to connect the assembly to stainless and disposable components; plus reliable regulatory, validation, and engineering support. “You can have the best filter in the world, but if you cannot help customers incorporate it into their process, you will not sell it,” says Mangeolle.

    Millipore’s latest filtration products include the Millistak+® HC Pod disposable depth filter, a line of fully disposable polyethersulfone membrane capsules, and the Integritest 4 Integrity Testing Instrument. Through its acquisition of Newport Biosystems, the company can also provide disposable process containers, tubing manifolds, and assembly systems.

    “The migration of vaccine manufacturing from eggs to cells has particularly benefited hollow fiber membrane filters,” says Ann O’Hara, who heads GE Healthcare’s (www.gehealthcare.com) bioprocess business. Hollow fiber units meet the needs for closed, aseptic systems that may be re-used or discarded after one run, depending on the processor’s requirements. According to O’Hara, hollow fiber membranes are easy on vaccines. “Since they move across the surface rather than being forced through, they don’t get beat up as much as with flat membranes,” says O’Hara

    A relatively new player in the filtration marketplace, GE Healthcare jump-started its business through the acquisition of AG Tech in 2001 and Novasep in 2002. GE built its filtration business on the successes of its chromatography systems, particularly its ÄKTA hardware platform. Powered by Unicorn Software (www.unicornsoftware.com), the ÄKTAcrossflow™ crossflow filtration product allows testing and optimization of crossflow ultrafiltration and microfiltration at small scale, which greatly facilitates scale-up. Many companies claim ease of scale-up with notoriously unpredictable molecules. “There will always be exceptions, and there are no guarantees,” says O’Hara, “but Unicorn at least allows you to apply science to scale-up by providing data all the way from bench to pilot and manufacturing scale.”

  • Efficiency vs. Flow

    The tradeoff between filtration efficiency and throughput will always be with us. Andrew Zydney, Ph.D., who heads the department of chemical engineering at the Pennsylvania State University, has discovered a way to diminish that compromise. Dr. Zydney uses electrically charged UF membranes that repel proteins of similar charge but simultaneously provide the high flux expected from filters with larger pores. Charged membranes are currently used industrially to recover electropaints, charged-particle pigments used in automobiles and appliances.

    Final product formulators will be interested in this charge-directed filtration since their product streams are relatively pure. Processors prefer to perform these purifications rapidly—in three hours or less—with as low a membrane area as possible, to avoid product degradation. “The normal way to achieve this is with tight membranes, which have low flux and require long processing times,” states Dr. Zydney.

    Charged ligands consist of quaternary amines and a variety of counter-ions. Dr. Zydney continues to investigate the impact of varying the ligands, charge density, length of the spacer arms connecting the charged ligand to the membrane material, and even the counter-ions. The standard negative ion for quaternary amines is hydroxide, but evidence from chromatography suggests the counter-ion may affect selectivity. “Whether this holds here or not remains to be seen,” says Dr. Zydney

    Dr. Zydney has been testing his invention in the real world through collaborations with Genentech (www.gene.com) and Millipore. “A commercial, first-generation product is in the works, but I don’t know the timeline,” he says

    Although some newer processes are shrinking as cell productivity and protein activity rises, existing processes, especially those for approved monoclonal antibodies, are growing in scale as the number of approved indications and market demand increase. For Mabs, that means bioreactors of 20,000 liters or more, this has created the need for higher-capacity and -flux culture filtration.

    The market is also shifting from the use of 0.2-micron filters to 0.1-micron to better protect against mycoplasma contaminations, according to Ralf Kuriyel, director of biopharm applications R&D at Pall (www.pall.com). In the absence of industry standards for 0.1 micron rated filters, the Parenteral Drug Association (www.pda.org) has formed a task force to establish standard challenge conditions for vendor qualification of mycoplasma retentive filters, which are nominally rated at 0.1 micron.

    In this particular case, the task force must balance the requirements of high throughput against retention of mycoplasma—essentially a safety tradeoff. Setting retention standards too high, for example at 100%, would dramatically degrade performance. The question is how much less than 100% retention of pathogenic bacteria will regulators accept?

    High titers, although always welcome, are somewhat of a mixed blessing since they present challenges for post-harvest clarification in the form of increased debris. The filtration following protein capture has also become more difficult due to the precipitation occurring during product pool neutralization. “Biotech manufacturers are looking for a single filter that has a high capacity for clean streams, such as buffers, and can also accommodate streams with higher solids loading,” says Kuriyel.

    Similarly, biomanufacturers are adopting small virus filtration early in bioprocesses, with a premium on high flux, capacity, and log removal values whose performance does not degrade as a function of throughput, even with challenging process fluids. “Users,” says Kuriyel, “prefer to conduct virus filtration at protein concentrations between 15–25 mg/mL, a target that may require the development of new membranes.”

    As the final protein concentrations reach 10 to 15 g/L, the final ultrafiltration/diafiltration step becomes difficult due to higher viscosities, necessitating design of products with reasonable feed channel pressure drops and adequate mass transfer properties. “Disposable TFF devices are attracting more interest for these applications,” says Kuriyel, “due to the simplicity of operation and validation.”

    In March, Pall introduced the Supor® UEKV and Fluorodyne® EX high-flow, sterilizing-grade cartridge filters. The 0.2-micron Supor UEKV combines Pall’s Ultipleat® membrane packing technology with a small-core design, providing a 50% increase in membrane area in the same size filter element. Supor UEKV also incorporates Pall’s machV polyethersulfone (PES) prefilter membrane layer with an asymmetric pore structure.

    Also utilizing the Ultipleat design, Fluorodyne EX features a built-in mach V PES prefilter over a high-flow 0.2-micron polyvinylidinedifluoride sterilizing membrane. The ability to steam sterilize both filters without pre-wetting reduces preparation time.

  • Specificity vs. Versatility

    “Filtration products are becoming more application-specific, designed to solve specific end user issues,” says Paul Priebe, who heads product management for process filtration products at Sartorius (www.sartorius.com). He gives the example of sterilizing-grade filters designed for buffer filtration, which in the past were selected for general performance. “But buffers require sterility assurance. You don’t have to worry about total throughput since the solutions are clean and will not clog. This fact allows filter optimization for high flow rate without regard to the reduced performance,” adds Priebe.

    The company’s Sartopore 2 HF is an example. Sartorius engineers removed the prefilter layer and added more filter surface area to give high flow rates, up to 40% higher than for general purpose filters, according to Sartorius, and reduce process times by up to 30%. Another example of filtration by design is the Sartorius ultrafiltration cassette, which is engineered for albumin concentration and diafiltration. “Again, in the past,” says Priebe, “we would have selected a generic 10 KD molecular weight cutoff cassette, which has a myriad of other uses and has been designed to offer good performance across those applications.”

    Although GEA Filtration, the membrane filtration division of Niro (www.niroinc.com), does not manufacture membranes, it claims to purchase and/or specify more of them than any process engineering membrane company in the U.S. After consulting with the customer, GEA will design and build spiral nanofiltration or ultrafiltration membrane systems.

    According to GEA Filtration’s biotech/pharma market manager, Bob Keefe, end-users increasingly demand filters that can do the work of multiple separation steps. For example, manufacturers of industrial enzymes have traditionally used centrifugation followed by rotary drum vacuum filtration in downstream operations. Today, they are likely to look for microfiltration to replace these two steps. “Ceramic microfiltration membranes will hold cellular debris 100% because it is a physical separation process based on particle size. You can’t even approach that with centrifugation,” states Keefe.

    From his perspective as global technology manager for DuPont’s (www.dupont.com) hybrid membrane technology unit, Tucker Norton sees a number of trends in the life science filtration industry: increasing regulatory demand for improved contaminant rejection and higher product purity; process simplification; the desire for higher throughput; and holding it all together, filter reliability and service. “Membranes and filters will continue to play an important role in addressing these trends,” he says.

    In response, DuPont has introduced the BarriRFlux™ line of liquid filtration products, with applications in bioprocessing as well as the food and beverage industries. BarriRFlux represents the first next-generation filtration technology from the company’s hybrid membrane group.

    Comprised of continuous, sub-micron fibers and available at commercial scale as membrane-like sheet structures, the new membranes are expected to fill the performance gap between traditional nonwovens and microporous films in critically clean life science applications by optimizing the balance between flux and retention.



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